Tumor cells have an absolute requirement for glutamine as a growth substrate. Glutamine is required as a precursor for both DNA synthesis and protein synthesis, and may also be used as a respiratory substrate. In experiments where glutamine metabolism in tumor cells has been specifically compared with that in non-transformed cells of the same origin, glutamine metabolism in the tumor cells has been found to be considerably faster. This is true for human hepatocytes and hepatoma cells (Souba, W., “Gutamine and Cancer,” Ann. Surg. 218:715-728 (1993)) and also for glutamine oxidation in rat kidney fibroblasts and rat fibrosarcoma cells (Fischer et al., “Adaptive Alterations in Cellular Metabolism and Malignant Transformation,” Ann. Surg. 227:627-634 (1998)).
The first reaction in glutamine metabolism is hydrolysis of glutamine to glutamate via the mitochondrial enzyme phosphate-dependent glutaminase. Two major isoforms of this enzyme have been characterized. These are known as the kidney form (K-type) which was first cloned from rat kidney (Shapiro et al., “Isolation, Characterisation, and In vitro Expression of a cDNA That Encodes the Kidney Isoenzyme of the Mitochondrial Glutaminase,” J. Biol. Chem. 266:18792-18796 (1991)) and is expressed in many mammalian tissues, and the liver form (L-type) (Chung-Bok et al., “Rat Hepatic Glutaminase, Identification of the Full Coding Sequence and Characterisation of a Functional Promoter,” Biochem. J. 324:193-200 (1997)) which was originally identified in post-natal liver. These two enzymes have different kinetic properties. Although the cDNAs encoding the two isoforms have regions of high sequence similarity, they also differ significantly elsewhere and the enzyme isoforms are the products of different genes (for a review see (Curthoys et al., “Regulation of Glutaminase Activity and Glutamine Metabolism,” Annu. Rev. Nutr. 16:133-159 (1995)). Glutamine metabolism is essential for tumor cell growth but there are few studies at present on glutaminase expression in tumor cells. In mouse Ehrlich ascites cells (Quesada et al., “Purification of Phosphate-Dependent Glutaminase from Isolated Mitochondria of Ehrlich Ascites-Tumor Cells,” Biochem. J. 255:1031-1035 (1988)) and rat fibrosarcoma cells (Fischer et al., “Adaptive Alterations in Cellular Metabolism and Malignant Transformation,” Ann. Surg. 227:627-634 (1998)), an enzyme with the kinetic properties of the K-type glutaminase is expressed. Rat and human hepatocytes express the L-type glutaminase, but this is not expressed in hepatoma cell lines, which express the K-type instead (Souba, W. W., “Glutamine and Cancer,” Ann. Surg. 218:715-728 (1993)). Inhibition of K-type glutaminase expression by anti-sense mRNA in Ehrlich ascites cells has been shown to decrease the growth and tumorigenicity of these cells (Lobo et al., “Inhibition of Glutaminase Expression by Antisense mRNA Decreases Growth and Tumorigenicity of Tumor Cells,” Biochem. J. 348:257-261 (2000)).
The present invention is directed to overcoming these and other deficiencies in the art.